A Clinical Case of Scrub Typhus in the United States Forces Korea Patient with Eschar and Genetic Identification of Orientia tsutsugamushi Using Multiplex PCR-Based Next-Generation Sequencing

Abstract

An epidemiological investigation was conducted for a scrub typhus case reported in a U.S. Forces Korea military patient in the Republic of Korea in November 2018. The patient had a fever, maculopapular rash, and an eschar. The full-length sequence of Orientia tsutsugamushi 56-kDa type-specific antigen (TSA) gene was obtained from eschar tissue by multiplex PCR-based Next Generation Sequencing for genetic identification. Based on the 56-kDa TSA gene, the O. tsutsugamushi aligned most closely with the Boryong strain.

Keywords: Orientia tsutsugamushi; genetic identification; scrub typhus; target enrichment next-generation sequencing.

Figure 1

An eschar on the lower left lower calf and maculopapular rash on the back of the USFK patient with Orientia tsutsugamushi infection. (A) Eschar location; (B) Back with maculopapular rash; (C) The location of Camp Humphreys and Jirisan National Park, Republic of Korea.

Figure 2

Phylogenetic analysis of the O. tsutsugamushi 56-kDa TSA gene identified from the USFK patient eschars. Phylogenetic relationships of the whole-genome sequences of O. tsutsugamushi 56-kDa TSA gene were inferred using Bayesian inference in BEAST (v1.10.4), using default priors and assuming homochromous tips. Upon running the Markov chain Monte Carlo analyses until adequate sample sizes (ESS > 200) were obtained, TreeAnnotator (v2.5.4) was used to summarize a maximum clade credibility tree from the posterior tree distribution, using a 10% burn-in. The Korean serotypes, Je-cheon (AF430143), Yeo-joo (AF430144), Pa-joo (AF430142), Young-worl (AF430141), Boryong (NC009488), and Yonchon (U19903), are indicated in bold. Topologies were evaluated by bootstrap analysis of 1000 iterations. Numbers along branches are bootstrap values. The scale bar represents nucleotide substitutions per site.